GENERATION OF CHROMOSOME FRAGMENT SPECIFIC BOVINE DNA-SEQUENCES BY MICRODISSECTION AND DOP-PCR

A rapid procedure for the defined isolation and characterization of si ngle bovine chromosome fragment specific probes is described. This has been developed as a technical pre requisite for the directed generati on of bovine DNA sequences. The specific regions 1q13-24, 5q21-24, 6q3 1-32, 7q21-22, 12q24-ter, and 20q12-ter of bovine GTG-banded metaphase chromosomes were microdissected and amplified by PCR with a degenerat e oligonucleotide primer and subsequently cloned into pBluescript II S K. The DNA probes generated were characterized by gel electrophoresis, dot blot analysis and rehybridization in situ to GTG-banded metaphase spreads. The position and size of the hybridization sites on the chro mosomes correspond exactly to the dissected chromosome areas and indic ate the complexity and specificity of the microdissected and amplified chromosome material.